Antigen recognition by T lymphocytes is mediated by diverse cell-surface glycoproteins known as T cell receptors (TCRs) which are composed of a and p (or y and 8) chains. In addition to antigenic peptides bound to MHC, a(3 TCRs interact with glycolipids presented by GDI and with a class of disease-causing molecules known as superantigens (SAGs). Although the specificity of y8 TCRs is less well understood, the major subset of human y8 T cells (Vy2/V52) recognizes small, aliphatic monoalkyl phosphate antigens from mycobacteria. Besides conventional cc(3 and y8 T cells, the T cell compartment includes natural killer (NK) T cells. NK T cells express NK inhibitory receptors that mediate self-recognition by binding MHC class I. These cells also express TCRs that recognize ceramide-containing glycolipids presented by GDI. Our aim is to elucidate the structural basis for three key aspects of antigen recognition by ocp, y5 and NK T cells: 1. Determination of the crystal structures of representative TCR |3 chain-SAG and SAG-MHC complexes in order to precisely define the diverse strategies that bacterial SAGs have evolved for binding TCR and MHC molecules. We previously determined the structures of a mouse TCR 3 chain (VP8.2) complexed with staphylococcal enterotoxins B and C3 (SEB, SEC3). We will now determine the structures of a human TCR P chain (V|32.1) complexed with toxic shock syndrome toxin-1 (TSST-1) and streptococcal pyrogenic exotoxin C (SPEC). We will also determine the structure of SEC3 complexed with HLA-DR1. This study will then be extended to SEA, SED and SPEC which, unlike SEC3, cross-link MHC molecules on APCs. 2. Determination of the structural basis for antigen recognition by human Vy2/V82 TCRs. We have recently solved the structure of the TCR VS domain and shown that it incorporates key structural elements of both antibody and CC0TCR V regions. We will now extend these studies to an associated Vy2/V82 TCR reactive with non-peptide prenyl pyrophosphates. 3. Determination of the structural basis for self-MHC and GDI recognition by NK T cells. We have crystallized a complex between the NK inhibitory receptor Ly-49A and its MHC class I ligand and a structure determination is underway.We will also express soluble forms of a canonical Vccl4-Ja281/Vp8.2 NK TCR for use in direct binding and co-crystallization experiments with specific CDl/glycolipid antigens.